Project Title: Nanolipoprotein supported multi-subunit vaccine to Chlamydia trachomatis Project Abstract Chlamydia trachomatis is the most common bacterial sexually transmitted infection (STI), affecting over 130 million people every year, and is the most common cause of preventable blindness worldwide. In the United States, STIs caused by C. trachomatis account for billions of dollars in annual costs (Gunn et al. 1998). Because the infection can be asymptomatic, it may go untreated for years and can result in long-term sequelae, including pelvic inflammatory disease, chronic abdominal pain, ectopic pregnancy, and infertility. Therefore, the pressing public health need for a vaccine to prevent diseases caused by C. trachomatis is clear. Despite considerable efforts to develop a chlamydial vaccine, none have been forthcoming. Studies have shown that immunization with the Chlamydia major outer membrane protein (MOMP) can induce significant protection against infection and disease in both mice and non-human primates if its native structure is preserved. However, formulation of MOMP vaccines is a major hurdle given that this protein has 16 transmembrane domains, is 40% hydrophobic, assembles as a homotrimer, and contains multiple cysteines that can form disulfide bridges. We have now demonstrated that we can produce a multi-oligomeric, SDS-resistant, and active form of MOMP using nanolipoprotein particles (NLPs). Furthermore, for the first time we can also produce full-length Chlamydia polymorphic membrane proteins (Pmps), another group of chlamydia surface antigens that have shown potent immunogenicity. The encoded proteins can be engineered to be serovar specific to make multi-serovar vaccines for mitigating affects associated with Chlamydia trachomatis pathogenicity. Finally, we have developed methods to combine MOMP with adjuvants to provide a unique vaccine formulation that was protective in mouse challenge studies. This initial breakthrough was achieved by combining synthetic biology approaches and cell-free co-expression of MOMP and PMPs with apolipoproteins. This proposal is focused on further extending MOMP presentation with associated PMPs, all formulated within NLPs, to achieve a highly protective vaccine against Chlamydia infections.